维药黑桑抑制酪氨酸酶活性部位的筛选及活性成分初步分析

Screening of Tyrosinase Inhibitory Parts and Analysis of Active Constituents from Uighur Medicine of Morus nigra

为了寻找天然、安全、高效的酪氨酸酶抑制剂,以维药黑桑茎枝为原料,通过乙醇提取和萃取分离,结合酪氨酸酶催化氧化左旋多巴(L-DOPA)速率法筛选酪氨酸酶抑制活性,获得主要活性部位;对主要活性部位进行HPLC指纹图谱表征和主要色谱峰指认。结果表明,黑桑茎枝乙醇粗提物(M)具有显著的酪氨酸酶抑制活性,IC_(50)达到0.785μg/ml,明显优于阳性对照曲酸(3.463μg/ml);M经萃取分离,得到主要活性部位二氯甲烷部位(M-2)和乙酸乙酯部位(M-3),活性均优于M,其中M-3的酪氨酸酶抑制活性最为显著,IC_(50)为0.036μg/ml,比曲酸强100倍,比M强约26倍。对活性部位M、M-2和M-3进行HPLC指纹图谱表征,通过色谱峰比较,指认了其中的4个色谱峰,均为桑根酮型黄酮。结果提示,维药黑桑是天然高效的酪氨酸酶抑制剂的良好来源,通过萃取分离,M中活性成分可得到有效富集。本研究为黑桑的进一步开发应用以及其活性物质基础阐明提供有益的参考。

In order to find natural, safe and highly efficient tyrosinase inhibitors, the stems of Morus nigra, a traditional Uighur medicine, were chosen as materials, and the active fractions were obtained by extraction and partition, combined with bioassay of tyrosinase inhibitory effects on the basis of evaluating oxidation rate of L-DOPA. Moreover, the active extracts were characterized by HPLC fingerprints in which the main chromatographic peaks were identified. The results exhibited that the ethanolic extract （M） showed potent tyrosinase inhibitory activity with IC50 of 0.785 μg/ml, obviously better than the positive control kojic acid （3.463 μg/ml）. After further extraction and separation, dichloromethane fraction （M-2） and ethyl acetate fraction （M-3） were obtained as the main active parts, with better tyrosinase inhibitory effects. The results showed that M-3 had an outstanding tyrosinase inhibitory effect with IC50 of 0.036 μg/ml, 100 times stronger than kojic acid, and 26 times stronger than M. The active parts of M, M-2 and M-3 were characterized by HPLC fingerprints, and 4 chromatographic peaks were identified as sanggenon-type flavanones. It was inferred that the plants ofMorus nigra might be a good source of natural tyrosinase inhibitors, and the active constituents could be effectively concentrated through extraction and separation. Our work might offer some references for the further development and application of Morus nigra as well as the elucidation of its bioactive constituents.